Dental plaque is an example of bacterial biofilm. Dental plaque is generally made up of bacteria and extracellular polymer substances (so called “EPS”). EPS are biopolymers of microbial origin in which biofilm microorganisms are embedded. The EPS Matrix: The “House of Biofilm Cells”, Hans-Curt Flemming, Thomas R. Neu and Daniel J. Wozniak J. Bacteriol. 2007, 189(22):7945. Dental plaque forms tartar and is associated with oral diseases such as caries and periodontal disease (e.g., gingivitis and periodontitis). Dental plaque can give rise to dental caries caused by the acid from the bacterial degradation of fermentable sugar. Therefore, dental plaque control and removal is important and the objective of many oral care compositions and regimens. Therefore, there is generally a continuing need to further understand the mechanisms of dental plaque formation and design oral care compositions that control or remove dental plaque.
Methods for quantitating the efficacy of oral care compositions (e.g., toothpaste, mouthwash etc.) at dislodging cells from biofilm test surfaces or inhibiting or delaying the accumulation of cells on a test surface have generally been described. Also, fluorescent probes, and confocal laser scanning microscopy (CLSM), have been generally used to assess ions and bacterial vitality of biofilm.
The role of ions in biofilm as well as use of certain ions in oral healthcare is a subject of ongoing study. Ions could inherently exist or be introduced in biofilm. For example, biofilm naturally contains calcium ions. There are abundant calcium ions (Ca2+) in dental plaque, and the amount of calcium in plaque is two to three times greater than in saliva. It is believed that positively charged calcium ions in the saliva mask the negative charges between bacteria and a tooth's surface allowing the bacteria to attach and multiply on the tooth surface, thus contributing to the formation of dental plaque. Accordingly, inhibiting the role of calcium in biofilm has been a target of therapeutic focus in some oral care compositions. Certain ions have been added to oral care compositions to help prevent dental plaque and oral disease. For example, cetylpyridinium chloride (CPC), stannous (Sn2+), silver (Ag+), copper (Cu2+) and zinc (Zn2+) are reported to help inhibit certain bacteria that can lead to tooth decay in human interproximal dental plaque. Fluoride (F−) is an ion found to help prevent caries.
It is reported that it is often difficult to purify EPS matrix constituent apart from other components of the biofilm such as cells. J. Bacteriol. 2007, 189(22):7945. However, there is a need for methods at quantitating ions localized in the EPS area of dental biofilm. For example, it is believed that calcium ions act as a “glue” or “scaffold” of EPS components. Other ions, such as stannous, zinc, fluoride could be released (or the release can be optimized) from oral care compositions to the EPS portion of dental biofilm to deliver increased efficacy. Inside cell clusters the locally high cell densities and the presence of EPS arrest the flow of water. Diffusion is limited in these biofilm systems because fluid flow is reduced and the diffusion distance is increased in the dental biofilm. As most pathogenic anaerobic microbes that cause gum problems typically reside in inner layer of EPS, these aforementioned ions need to diffuse deeper to treat these microbes. Accordingly there is a need for methods to study the diffusion pattern of these ions in the EPS area of dental biofilm.